A novel, rapidly acquired and persistent spatial memory task that induces immediate early gene expression.

BACKGROUND: The Morris water maze task is a hippocampus-dependent learning and memory test that typically takes between 3 days to 2 weeks of training. This task is used to assess spatial learning and induces the expression of genes known to be crucial to learning and memory in the hippocampus. A major caveat in the protocol is the prolonged duration of training, and difficulty of assessing the time during training in which animals have learned the task. We introduce here a condensed version of the task that like traditional water maze tasks, creates lasting hippocampus-dependent spatial cognitive maps and elicits gene expression following learning. METHODS: This paradigm was designed for rats to quickly acquire a hippocampus-dependent spatial cognitive map and retain this memory for at least 24 hours. To accomplish this, we interspersed visible and hidden training trials, delivering them in a massed fashion so training takes a maximum of 15 minutes. Learning was assessed based on latencies to the platform during each training trial, as well as time spent in the goal quadrant during probe testing 30 minutes and 24 hours after training. Normal rats were compared to two impaired cohorts (rats with fimbria-fornix lesions and rats administered NMDA receptor antagonist (CPP)). To quantitate hippocampal expression of known learning genes, real-time polymerase chain reaction (RT-PCR) was performed on hippocampal cDNA. RESULTS: We show that massed training using alternating visible and hidden training trials generates robust short-term working and long-term reference memories in rats. Like the traditional Morris water maze paradigm, this task requires proper hippocampal function, as rats with fimbria-fornix lesions and rats administered CPP fail to learn the spatial component of the task. Furthermore, training in this paradigm elicits hippocampal expression of genes upregulated following learning in a variety of spatial tasks: homer1a, cfos and zif268. CONCLUSIONS: We introduce here a condensed version of the Morris water maze, which is like a traditional water maze paradigm, in that it is hippocampus-dependent, and elicits hippocampal expression of learning genes. However, this task is administered in 15 minutes and induces spatial memory for at least 24 hours.

Connexin43 Expression and Associated Chronic Inflammation Presages the Development of Cerebral Radiation Necrosis.

Cerebral radiation necrosis (CRN) is a delayed complication of radiosurgery that can result in severe neurological deficits. The biological changes leading to necrotic damage may identify therapeutic targets for this complication. Connexin43 expression associated with chronic inflammation may presage the development of CRN. A mouse model of delayed CRN was used. The left hemispheres of adult female mice were irradiated with single-fraction, high-dose radiation using a Leksell Gamma Knife. The brains were collected 1 and 4 days, and 1-3 weeks after the radiation. The expression of connexin43, interleukin-1ß (IL-1ß), GFAP, isolectin B-4, and fibrinogen was evaluated using immunohistochemical staining and image analysis. Compared with the baseline, the area of connexin43 and IL-1ß staining was increased in ipsilateral hemispheres 4 days after radiation. Over the following 3 weeks, the density of connexin43 gradually increased in parallel with progressive increases in GFAP, isolectin B-4, and fibrinogen labeling. The overexpression of connexin43 in parallel with IL-1ß spread into the affected brain regions first. Further intensified upregulation of connexin43 was associated with escalated astrocytosis, microgliosis, and blood-brain barrier breach. Connexin43-mediated inflammation may underlie radiation necrosis and further investigation of connexin43 hemichannel blockage is merited for the treatment of CRN.

Perfluorocarbon emulsions radiosensitise brain tumors in carbogen breathing mice with orthotopic GL261 gliomas.

BACKGROUND: Tumour hypoxia limits the effectiveness of radiation therapy. Delivering normobaric or hyperbaric oxygen therapy elevates pO2 in both tumour and normal brain tissue. However, pO2 levels return to baseline within 15 minutes of stopping therapy. AIM: To investigate the effect of perfluorocarbon (PFC) emulsions on hypoxia in subcutaneous and intracranial mouse gliomas and their radiosensitising effect in orthotopic gliomas in mice breathing carbogen (95%O2 and 5%CO2). RESULTS: PFC emulsions completely abrogated hypoxia in both subcutaneous and intracranial GL261 models and conferred a significant survival advantage orthotopically (Mantel Cox: p = 0.048) in carbogen breathing mice injected intravenously (IV) with PFC emulsions before radiation versus mice receiving radiation alone. Carbogen alone decreased hypoxia levels substantially and conferred a smaller but not statistically significant survival advantage over and above radiation alone. CONCLUSION: IV injections of PFC emulsions followed by 1h carbogen breathing, radiosensitises GL261 intracranial tumors.

Enhanced immunity to intracerebral breast cancer in mice immunized with a cDNA-based vaccine enriched for immunotherapeutic cells.

Structural differences between malignant and nonmalignant cells of the same individual form the basis of clinical immunotherapeutic strategies. Previously, we reported the therapeutic properties of a vaccine prepared by transfer of a cDNA-expression library from breast cancer cells into a highly immunogenic allogeneic fibroblast cell line where genes specifying an array of breast cancer antigens were expressed. Here, we report the application of this cell-based vaccination strategy for breast cancer metastatic to the brain. As relatively few cells in the vaccine were expected to have incorporated cDNA fragments specifying breast cancer antigens (most specify normal cellular constituents), an enrichment strategy was employed to increase the proportion of immunotherapeutic cells. Enhanced immunity to the neoplasm mediated predominantly by CD4+, CD8+, and NK/LAK cells was generated in the spleens of mice injected intracerebrally into the tumor bed with cells from the enriched vaccine, which translated into prolonged survival. Regulatory T cells (CD4+CD25+Foxp3+-positive) were relatively deficient in the spleen cells from tumor-bearing mice injected intracerebrally with the enriched vaccine.